Intraventricular enzyme delivery for lysosomal storage diseases

ABSTRACT

Lysosomal storage diseases can be successfully treated using intraventricular delivery of the enzyme which is etiologically deficient in the disease. The administration can be performed slowly to achieve maximum effect. Surprisingly, effects are seen on both sides of the blood-brain barrier, making this an ideal delivery means for lysosomal storage diseases which affect both brain and visceral organs.

TECHNICAL FIELD OF THE INVENTION

This invention is related to the area of lysosomal storage diseases. Inparticular, it relates to the treatment and/or prevention of thesediseases by enzyme replacement therapy.

SUMMARY OF THE INVENTION

A group of metabolic disorders known as lysosomal storage diseases(LSDs) includes over forty genetic disorders, many of which involvegenetic defects in various lysosomal hydrolases. Representativelysosomal storage diseases and the associated defective enzymes arelisted in Table 1.

TABLE 1 Lysosomal storage disease Defective enzymeAspartylglucosaminuria Aspartylglucosaminidase Fabryalpha.-Galactosidase A Infantile Batten Disease* Palmitoyl ProteinThioesterase (CNL1) Classic Late Infantile Batten Tripeptidyl PeptidaseDisease* (CNL2) Juvenile Batten Disease* Lysosomal Transmembrane Protein(CNL3) Batten, other forms* (CNL4-CNL8) Multiple gene productsCystinosis Cysteine transporter Farber Acid ceramidase Fucosidosis Acid.alpha.-L-fucosidase Galactosidosialidosis Protective protein/cathepsinA Gaucher types 1, 2*, and 3* Acid .beta.-glucosidase, or G.sub.M1gangliosidosis* Acid .beta.-galactosidase Hunter* Iduronate-2-sulfataseHurler-Scheie* alpha.-L-Iduronidase Krabbe* Galactocerebrosidase.alpha.-Mannosidosis* Acid .alpha.-mannosidase. beta.-Mannosidosis* Acid.beta.-mannosidase Maroteaux-Lamy Arylsulfatase B Metachromaticleukodystrophy* Arylsulfatase A Morquio AN-Acetylgalactosamine-6-sulfate Morquio B Acid .beta.-galactosidaseMucolipidosis II/III* N-Acetylglucosamine-1- Niemann-Pick A*, B Acidsphingomyelinase Niemann-Pick C* NPC-1 Pompe* Acid .alpha.-glucosidaseSandhoff* .beta.-Hexosaminidase B Sanfilippo A* Heparan N-sulfataseSanfilippo B* .alpha.-N-Acetylglucosaminidase Sanfilippo C* Acetyl-CoA:alpha.-glucosaminide Sanfilippo D* N-Acetylglucosamine-6-sulfateSchindler Disease* .alpha.-N-Acetylgalactosaminidase Schindler-Kanzaki.alpha.-N-Acetylgalactosaminidase Sialidosis .alpha.-Neuramidase Sly*.beta.-Glucuronidase Tay-Sachs* .beta.-Hexosaminidase A Wolman* AcidLipase *CNS involvement

The hallmark feature of LSDs is the abnormal accumulation of metabolitesin the lysosomes which leads to the formation of large numbers ofdistended lysosomes in the perikaryon. A major challenge to treatingLSDs (as opposed to treating an organ specific enzymopathy, e.g., aliver-specific enzymopathy) is the need to reverse lysosomal storagepathology in multiple separate tissues. Some LSDs can be effectivelytreated by intravenous infusion of the missing enzyme, known as enzymereplacement therapy (ERT). For example, Gaucher type 1 patients haveonly visceral disease and respond favorably to ERT with recombinantglucocerebrosidase (Cerezyme™, Genzyme Corp.). However, patients withmetabolic disease that affects the CNS (e.g., type 2 or 3 Gaucherdisease) only respond partially to intravenous ERT because thereplacement enzyme is prevented from entering the brain by the bloodbrain barrier (BBB). Furthermore, attempts to introduce a replacementenzyme into the brain by direct injection have been limited in part dueto enzyme cytotoxicity at high local concentrations and limitedparenchymal diffusion rates in the brain (Partridge, Peptide DrugDelivery to the Brain, Raven Press, 1991).

One exemplary LSD, is Niemann-Pick disease type A (NPA). According toUniProtKB/Swiss-Prot entry P17405, defects in the SMPD1 gene, located onchromosome 11, (11p15.4-p15.1), are the cause of Niemann-Pick diseasetype A (NPA), also referred to as the classical infantile form of thedisease. Niemann-Pick disease is a clinically and geneticallyheterogeneous recessive disorder. It is caused by the accumulation ofsphingomyelin and other metabolically related lipids in the lysosomes,resulting in neurodegeneration starting from early life. Patients mayshow xanthomas, pigmentation, hepatosplenomegaly, lymphadenopathy andmental retardation. Niemann-Pick disease occurs more frequently amongindividuals of Ashkenazi Jewish ancestry than in the general population.NPA is characterized by very early onset in infancy and a rapidlyprogressive course leading to death by three years. The acidsphingomyelinase enzyme (ASM) that is defective in NPA convertssphingomyelin to ceramide. ASM also has phospholipase C activitiestoward 1,2-diacylglycerolphosphocholine and1,2-diacylglycerolphosphoglycerol. The enzyme converts

Sphingomyelin+H₂O→N-acylsphingosine+choline phosphate.

In accordance with the present invention, lysosomal storage diseasessuch as any of the diseases identified in Table 1 above, e.g.,Niemann-Pick disease type A or B, are treated and/or prevented usingintraventricular delivery to the brain of the enzyme which isetiologically deficient in the disease. The administration can beperformed slowly to achieve maximum effect. Effects are seen on bothsides of the blood-brain barrier, making this a useful delivery meansfor lysosomal storage diseases which affect the brain and/or visceralorgans. In a first aspect, the invention thus provides for a method oftreating or preventing a lysosomal storage disease in patient, whichdisease is caused by an enzyme deficiency, the method comprisingadministering the enzyme to the patient via intraventricular delivery tothe brain. In a related aspect, the invention provides for the use of anenzyme for the manufacture of a medicament for the treatment orprevention of a lysosomal storage disease in a patient, which disease iscaused by a deficiency of the enzyme in the patient, wherein thetreatment or prevention comprises the intraventricular administration ofthe enzyme to the brain. The enzyme deficiency may be caused by e.g., adefect in the expression of the enzyme or by a mutation in the enzymewhich leads to a reduced level of activity (e.g., the enzyme beinginactive) or an increased rate of clearance/breakdown of the enzyme invivo. The deficiency may lead to an accumulation of an enzyme'ssubstrate and the administration of the enzyme may lead to a reductionin the level of the substrate in the brain. The lysosomal storagedisease may be any of the diseases identified in Table 1 above. Theenzyme may be a lysosomal hydrolase.

According to one embodiment of the invention, a patient withNiemann-Pick A or B disease is treated. An acid sphingomyelinase isadministered to the patient via intraventricular delivery to the brainin an amount sufficient to reduce sphingomyelin levels in said brain.

Another aspect of the invention is a kit for treating or preventing alysosomal storage disease in patient, which disease is caused by anenzyme deficiency. The kit comprises the enzyme that is deficient, and acatheter and/or pump for delivery of the enzyme to one or moreventricles in the brain. The catheter and/or pump may be specificallydesigned and/or adapted for intraventricular delivery. According to oneembodiment, the invention provides a kit for treating a patient withNiemann-Pick A or B disease. The kit comprises an acid sphingomyelinase,and a catheter for delivery of said acid sphingomyelinase to thepatient's brain ventricles.

Yet another aspect of the invention is a kit for treating a patient withNiemann-Pick A or B disease. The kit comprises an acid sphingomyelinaseand a pump for delivery of said acid sphingomyelinase to the patient'sbrain ventricles.

According to the invention, a patient can be treated who has a lysosomalstorage disease which is caused by an enzyme deficiency which leads toaccumulation of the enzyme's substrate. Such diseases include Gaucherdisease, MPS I and II disease, Pompe disease, and Batten disease (CLN2),among others. The enzyme defective in the particular disease isadministered to the patient via intraventricular delivery to the brain.It may be administered to the lateral ventricles and/or to the fourthventricle. The rate of administration of the enzyme in accordance withthe present invention is such that the administration of a single dosemay be as a bolus or may take about 1-5 minutes, about 5-10 minutes,about 10-30 minutes, about 30-60 minutes, about 1-4 hours, or consumesmore than four, five, six, seven, or eight hours. Substrate levels insaid brain may thereby be reduced. The administration of a single dosemay take more than 1 minute, more than 2 minutes, more than 5 minutes,more than 10 minutes, more than 20 minutes, more than 30 minutes, morethan 1 hour, more than 2 hours, or more than 3 hours.

These and other embodiments which will be apparent to those of skill inthe art upon reading the specification provide the art with methods andkits for the treatment and/or prevention of lysosomal storage diseases,in particular those involving both CNS and visceral pathologies.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows diagram of sections of brain that were analyzed forsphingomyelin. S1 is at the front of brain and S5 is at the back ofbrain.

FIG. 2 shows that intraventricular administration of rhASM reduces SPMlevels in the ASMKO mouse brain.

FIGS. 3A, 3B, and 3C shows intraventricular administration of rhASMreduces SPM levels in the ASMKO liver (FIG. 3A), spleen (FIG. 3C), andlung (FIG. 3B).

FIG. 4 shows hASM staining in the brain following intraventricularinfusion.

FIG. 5 shows that intraventricular infusion of rhASM over a 6 h periodreduces SPM levels in the ASMKO mouse brain.

FIGS. 6A, 6B, and 6C shows that intraventricular infusion of rhASM overa 6 h period reduces SPM levels in ASMKO liver (FIG. 6A), serum (FIG.6C), and lung (FIG. 6B).

FIG. 7 shows documented hASM variants and their relationship to diseaseor enzyme activity.

FIG. 8 shows a cross section view of the brain with the ventriclesindicated.

FIGS. 9A and 9B show lateral and superior views, respectively, of theventricles.

FIG. 10 shows injection into the lateral ventricles.

DETAILED DESCRIPTION OF THE INVENTION

The practice of the present invention will employ, unless otherwiseindicated, conventional techniques of immunology, molecular biology,microbiology, cell biology and recombinant DNA, which are within theskill of the art. See, e.g., Sambrook, Fritsch and Maniatis, MOLECULARCLONING: A LABORATORY MANUAL, 2^(nd) edition (1989); CURRENT PROTOCOLSIN MOLECULAR BIOLOGY (F. M. Ausubel, et al. eds., (1987)); the seriesMETHODS IN ENZYMOLOGY (Academic Press, Inc.): PCR 2: A PRACTICALAPPROACH (M. J. MacPherson, B. D. Hames and G. R. Taylor eds. (1995)),Harlow and Lane, eds. (1988) ANTIBODIES, A LABORATORY MANUAL, and ANIMALCELL CULTURE (R. I. Freshney, ed. (1987)).

As used in the specification and claims, the singular forms “a,” “an”and “the” include plural references unless the context clearly dictatesotherwise. For example, the term “a cell” includes a plurality of cells,including mixtures thereof.

As used herein, the term “comprising” is intended to mean that thecompositions and methods include the recited elements, but not excludingothers. “Consisting essentially of” when used to define compositions andmethods, shall mean excluding other elements of any essentialsignificance to the combination. Thus, a composition consistingessentially of the elements as defined herein would not exclude tracecontaminants from the isolation and purification method andpharmaceutically acceptable carriers, such as phosphate buffered saline,preservatives, and the like. “Consisting of” shall mean excluding morethan trace elements of other ingredients and excluding substantialmethod steps for administering the compositions or medicaments inaccordance with this invention. Embodiments defined by each of thesetransition terms are within the scope of this invention.

All numerical designations, e.g., pH, temperature, time, concentration,and molecular weight, including ranges, are approximations which arevaried (+) or (−) by increments of 0.1. It is to be understood, althoughnot always explicitly stated that all numerical designations arepreceded by the term “about.” It also is to be understood, although notalways explicitly stated, that the reagents described herein are merelyexemplary and that equivalents of such that are known in the art mayalso be used.

The terms “therapeutic,” “therapeutically effective amount,” and theircognates refer to that amount of a substance, e.g., enzyme or protein,that results in prevention or delay of onset, or amelioration, of one ormore symptoms of a disease in a subject, or an attainment of a desiredbiological outcome, such as correction of neuropathology. The term“therapeutic correction” refers to that degree of correction whichresults in prevention or delay of onset, or amelioration, of one or moresymptoms in a subject. The effective amount can be determined by knownempirical methods.

A “composition” or “medicament” is intended to encompass a combinationof an active agent, e.g., an enzyme, and a carrier or other material,e.g., a compound or composition, which is inert (for example, adetectable agent or label) or active, such as an adjuvant, diluent,binder, stabilizer, buffer, salt, lipophilic solvent, preservative,adjuvant or the like, or a mixture of two or more of these substances.Carriers are preferably pharmaceutically acceptable. They may includepharmaceutical excipients and additives, proteins, peptides, aminoacids, lipids, and carbohydrates (e.g., sugars, includingmonosaccharides, di-, tri-, tetra-, and oligosaccharides; derivatizedsugars such as alditols, aldonic acids, esterified sugars and the like;and polysaccharides or sugar polymers), which can be present singly orin combination, comprising alone or in combination 1-99.99% by weight orvolume. Exemplary protein excipients include serum albumin such as humanserum albumin (HSA), recombinant human albumin (rHA), gelatin, casein,and the like. Representative amino acid/antibody components, which canalso function in a buffering capacity, include alanine, glycine,arginine, betaine, histidine, glutamic acid, aspartic acid, cysteine,lysine, leucine, isoleucine, valine, methionine, phenylalanine,aspartame, and the like. Carbohydrate excipients are also intendedwithin the scope of this invention, examples of which include but arenot limited to monosaccharides such as fructose, maltose, galactose,glucose, D-mannose, sorbose, and the like; disaccharides, such aslactose, sucrose, trehalose, cellobiose, and the like; polysaccharides,such as raffinose, melezitose, maltodextrins, dextrans, starches, andthe like; and alditols, such as mannitol, xylitol, maltitol, lactitol,xylitol sorbitol (glucitol) and myoinositol.

The term carrier also includes a buffer or a pH adjusting agent or acomposition containing the same; typically, the buffer is a saltprepared from an organic acid or base. Representative buffers includeorganic acid salts such as salts of citric acid, ascorbic acid, gluconicacid, carbonic acid, tartaric acid, succinic acid, acetic acid, orphthalic acid, Tris, tromethamine hydrochloride, or phosphate buffers.Additional carriers include polymeric excipients/additives such aspolyvinylpyrrolidones, ficolls (a polymeric sugar), dextrates (e.g.,cyclodextrins, such as 2-hydroxypropyl-.quadrature.-cyclodextrin),polyethylene glycols, flavoring agents, antimicrobial agents,sweeteners, antioxidants, antistatic agents, surfactants (e.g.,polysorbates such as “TWEEN 20” and “TWEEN 80”), lipids (e.g.,phospholipids, fatty acids), steroids (e.g., cholesterol), and chelatingagents (e.g., EDTA).

As used herein, the term “pharmaceutically acceptable carrier”encompasses any of the standard pharmaceutical carriers, such as aphosphate buffered saline solution, water, and emulsions, such as anoil/water or water/oil emulsion, and various types of wetting agents.The compositions and medicaments which are manufactured and/or used inaccordance with the present invention and which include the particularenzyme whose deficiency is to be corrected can include stabilizers andpreservatives and any of the above noted carriers with the additionalproviso that they be acceptable for use in vivo. For examples ofcarriers, stabilizers and adjuvants, see Martin REMINGTON'S PHARM. SCI.,15th Ed. (Mack Publ. Co., Easton (1975) and Williams & Williams, (1995),and in the “PHYSICIAN'S DESK REFERENCE”, 52^(nd) ed., Medical Economics,Montvale, N.J. (1998).

A “subject,” “individual” or “patient” is used interchangeably herein,which refers to a vertebrate, preferably a mammal, more preferably ahuman. Mammals include, but are not limited to, mice, rats, monkeys,humans, farm animals, sport animals, and pets.

As used herein, the term “modulate” means to vary the amount orintensity of an effect or outcome, e.g., to enhance, augment, diminishor reduce.

As used herein the term “ameliorate” is synonymous with “alleviate” andmeans to reduce or lighten. For example, one may ameliorate the symptomsof a disease or disorder by making them more bearable.

For identification of structures in the human brain, see, e.g., TheHuman Brain: Surface, Three-Dimensional Sectional Anatomy With MRI, andBlood Supply, 2nd ed., eds. Deuteron et al., Springer Vela, 1999; Atlasof the Human Brain, eds. Mai et al., Academic Press; 1997; and Co-PlanarStereotaxic Atlas of the Human Brain: 3-Dimensional Proportional System:An Approach to Cerebral Imaging, eds. Tamarack et al., Thyme MedicalPub., 1988. For identification of structures in the mouse brain, see,e.g., The Mouse Brain in Stereotaxic Coordinates, 2nd ed., AcademicPress, 2000.

The inventors have discovered that intraventricular delivery to thebrain of lysosomal hydrolase enzymes to patients who are deficient inthe enzymes, leads to improved metabolic status of both the brain andthe affected visceral (non-CNS) organs. This is particularly true whenthe delivery rate is slow, relative to a bolus delivery. Lysosomalstorage diseases caused by a deficiency in a particular enzyme, such asthose diseases identified in Table 1 above, may therefore be treated orprevented by the intraventricular administration of the respectiveenzyme. One particularly useful enzyme for treating Niemann-Pick A or Bis acid sphingomyelinase (ASM), such as that shown in SEQ ID NO: 1.¹ Oneparticularly useful enzyme for treating Gaucher disease isglucocerebrosidase. One particularly useful enzyme for treating MPS Idisease is alpha-L-Iduronidase. One particularly useful enzyme fortreating MPS II disease is iduronate-2-sulfatase. One particularlyuseful enzyme for treating Pompe disease, or glycogen storage diseasetype II (GSDII), also termed acid maltase deficiency (AMD) is acidalpha-glucosidase. One particularly useful enzyme for treating classiclate infantile Batten disease (CLN2) is tripeptidyl peptidase. Theenzymes that are used and/or administered in accordance with the presentinvention may be recombinant forms of the enzymes which are producedusing methods that are well-known in the art. In one embodiment, theenzyme is a recombinant human enzyme. Residues 1-46 constitute thesignal sequence which is cleaved upon secretion.

Administration of lysosomal enzymes, and more particularly lysosomalhydrolase enzymes, to patients who are deficient in the enzymes may beperformed into any one or more of the brain's ventricles, which arefilled with cerebrospinal fluid (CSF). CSF is a clear fluid that fillsthe ventricles, is present in the subarachnoid space, and surrounds thebrain and spinal cord. CSF is produced by the choroid plexuses and viathe weeping or transmission of tissue fluid by the brain into theventricles. The choroid plexus is a structure lining the floor of thelateral ventricle and the roof of the third and fourth ventricles.Certain studies have indicated that these structures are capable ofproducing 400-600 ccs of fluid per day consistent with an amount to fillthe central nervous system spaces four times in a day. In adults, thevolume of this fluid has been calculated to be from 125 to 150 ml (4-5oz). The CSF is in continuous formation, circulation and absorption.Certain studies have indicated that approximately 430 to 450 ml (nearly2 cups) of CSF may be produced every day. Certain calculations estimatethat production equals approximately 0.35 ml per minute in adults and0.15 per minute in infants. The choroid plexuses of the lateralventricles produce the majority of CSF. It flows through the foramina ofMonro into the third ventricle where it is added to by production fromthe third ventricle and continues down through the aqueduct of Sylviusto the fourth ventricle. The fourth ventricle adds more CSF; the fluidthen travels into the subarachnoid space through the foramina ofMagendie and Luschka. It then circulates throughout the base of thebrain, down around the spinal cord and upward over the cerebralhemispheres. The CSF empties into the blood via the arachnoid villi andintracranial vascular sinuses, thereby delivering the enzymes that areinfused into the ventricles to not only the brain but also to thevisceral organs that are known to be affected in LSDs.

Although a particular amino acid sequence is shown in SEQ ID NO: 1,variants of that sequence which retain activity, e.g., normal variantsin the human population, can be used as well. Typically these normalvariants differ by just one or two residues from the sequence shown inSEQ ID NO: 1. The variants of SEQ ID NO: 1 that are to be used inaccordance with the present invention, whether naturally occurring ornot, should be at least 95%, 96%, 97%, 98%, or 99% identical to SEQ IDNO: 1. Variants of other enzymes may be used in accordance with thepresent invention. However, irrespective of the enzyme that is used,variants which are associated with disease or reduced activity shouldnot be used. Typically the mature form of the enzyme will be delivered.In the case of SEQ ID NO: 1, the mature form will begin with residue 47as shown in SEQ ID NO: 1. Variants which are associated with disease areshown in FIG. 7. In a similar manner, normal variants in the humanpopulation of such LSD enzymes as glucocerebrosidase,alpha-L-Iduronidase, iduronate-2-sulfatase, acid alpha-glucosidase, andtripeptidyl peptidase that which retain enzymatic activity can be usedas well.

Kits according to the present invention are assemblages of separatecomponents. While they can be packaged in a single container, they canbe subpackaged separately. Even a single container can be divided intocompartments. Typically a set of instructions will accompany the kit andprovide instructions for delivering the enzymes, e.g., the lysosomalhydrolase enzymes, intraventricularly. The instructions may be inprinted form, in electronic form, as an instructional video or DVD, on acompact disc, on a floppy disc, on the internet with an address providedin the package, or a combination of these means. Other components, suchas diluents, buffers, solvents, tape, screws, and maintenance tools canbe provided in addition to the enzyme, one or more cannulae orcatheters, and/or a pump.

The populations treated by the methods of the invention include, but arenot limited to, patients having, or who are at risk for developing, aneurometabolic disorder, e.g., an LSD, such as the diseases listed inTable 1, particularly if such a disease affects the CNS and visceralorgans. In an illustrative embodiment, the disease is type ANiemann-Pick disease.

An ASM or other lysosomal hydrolase enzyme can be incorporated into apharmaceutical composition useful to treat, e.g., inhibit, attenuate,prevent, or ameliorate, a condition characterized by an insufficientlevel of a lysosomal hydrolase activity. The pharmaceutical compositionwill be administered to a subject suffering from a lysosomal hydrolasedeficiency or someone who is at risk of developing said deficiency. Thecompositions should contain a therapeutic or prophylactic amount of theASM or other lysosomal hydrolase enzyme, in apharmaceutically-acceptable carrier. The pharmaceutical carrier can beany compatible, non-toxic substance suitable to deliver the polypeptidesto the patient. Sterile water, alcohol, fats, and waxes may be used asthe carrier. Pharmaceutically-acceptable adjuvants, buffering agents,dispersing agents, and the like, may also be incorporated into thepharmaceutical compositions. The carrier can be combined with the ASM orother lysosomal hydrolase enzyme in any form suitable for administrationby intraventricular injection or infusion (which form is also possiblysuitable for intravenous or intrathecal administration) or otherwise.Suitable carriers include, for example, physiological saline,bacteriostatic water, Cremophor EL™ (BASF, Parsippany, N.J.) orphosphate buffered saline (PBS), other saline solutions, dextrosesolutions, glycerol solutions, water and oils emulsions such as thosemade with oils of petroleum, animal, vegetable, or synthetic origin(peanut oil, soybean oil, mineral oil, or sesame oil). An artificial CSFcan be used as a carrier. The carrier will preferably be sterile andfree of pyrogens. The concentration of the ASM or other lysosomalhydrolase enzyme in the pharmaceutical composition can vary widely,i.e., from at least about 0.01% by weight, to 0.1% by weight, to about1% weight, to as much as 20% by weight or more of the total composition.

For intraventricular administration of ASM or other lysosomal hydrolaseenzyme, the composition must be sterile and should be fluid. It must bestable under the conditions of manufacture and storage and must bepreserved against the contaminating action of microorganisms such asbacteria and fungi. Prevention of the action of microorganisms can beachieved by various antibacterial and antifungal agents, for example,parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and thelike. In many cases, it will be preferable to include isotonic agents,for example, sugars, polyalcohols such as mannitol, sorbitol, sodiumchloride in the composition.

Dosage of ASM or other lysosomal hydrolase enzyme may vary somewhat fromindividual to individual, depending on the particular enzyme and itsspecific in vivo activity, the route of administration, the medicalcondition, age, weight or sex of the patient, the patient'ssensitivities to the ASM or other lysosomal hydrolase enzyme orcomponents of the vehicle, and other factors which the attendingphysician will be capable of readily taking into account. While dosagesmay vary depending on the disease and the patient, the enzyme isgenerally administered to the patient in amounts of from about 0.1 toabout 1000 milligrams per 50 kg of patient per month. In one embodiment,the enzyme is administered to the patient in amounts of about 1 to about500 milligrams per 50 kg of patient per month. In other embodiments, theenzyme is administered to the patient in amounts of about 5 to about 300milligrams per 50 kg of patient per month, or about 10 to about 200milligrams per 50 kg of patient per month.

The rate of administration is such that the administration of a singledose may be administered as a bolus. A single dose may also be infusedover about 1-5 minutes, about 5-10 minutes, about 10-30 minutes, about30-60 minutes, about 1-4 hours, or consumes more than four, five, six,seven, or eight hours. It may take more than 1 minute, more than 2minutes, more than 5 minutes, more than 10 minutes, more than 20minutes, more than 30 minutes, more than 1 hour, more than 2 hours, ormore than 3 hours. Applicants have observed that, while bolusintraventricular administration may be effective, slow infusion is veryeffective. While applicants do not wish to be bound by any particulartheory of operation, it is believed that the slow infusion is moreeffective due to the turn-over of the cerebrospinal fluid (CSF). Whileestimates and calculations in the literature vary, the cerebrospinalfluid is believed to turn over within about 4, 5, 6, 7, or 8 hours inhumans. In one embodiment, the slow infusion time of the inventionshould be metered so that it is about equal to or greater than theturn-over time of the CSF. Turn-over time may depend on the species,size, and age of the subject but may be determined using methods knownin the art. Infusion may also be continuous over a period of one or moredays. The patient may be treated once, twice, or three or more times amonth, e.g., weekly, e.g. every two weeks. Infusions may be repeatedover the course of a subject's life as dictated by re-accumulation ofthe disease's substrate in the brain or visceral organs. Re-accumulationmay be determined by any of the techniques that are well known in theart for the identification and quantization of the relevant substrate,which techniques may be performed on one or more samples taken from thebrain and/or from one or more of the visceral organs. Such techniquesinclude enzymatic assays and/or immunoassays, e.g. radioimmunoassays orELISAs.

The CSF empties into the blood via the arachnoid villi and intracranialvascular sinuses, thereby delivering the infused enzymes to the visceralorgans that are known to be affected in LSDs. The visceral organs whichare often affected in Niemann-Pick disease are the lungs, spleen,kidney, and liver. The slow intraventricular infusion providesdiminished amounts of the substrate for an administered enzyme in atleast the brain and potentially in visceral organs. The reduction insubstrate accumulated in the brain, lungs, spleen, kidney, and/or livermay be dramatic. Reductions of greater that 10%, 20%, 30%, 40%, 50%,60%, 70%, 80%, 90% can be achieved. The reduction achieved is notnecessarily uniform from patient to patient or even from organ to organwithin a single patient. Reductions can be determined by any of thetechniques that are well known in the art, e.g., by enzymatic assaysand/or immunoassay techniques, as discussed elsewhere herein.

If desired, the human brain structure can be correlated to similarstructures in the brain of another mammal. For example, most mammals,including humans and rodents, show a similar topographical organizationof the entorhinal-hippocampus projections, with neurons in the lateralpart of both the lateral and medial entorhinal cortex projecting to thedorsal part or septal pole of the hippocampus, whereas the projection tothe ventral hippocampus originates primarily from neurons in medialparts of the entorhinal cortex (Principles of Neural Science, 4th ed.,eds Kandel et al., McGraw-Hill, 1991; The Rat Nervous System, 2nd ed.,ed. Paxinos, Academic Press, 1995). Furthermore, layer II cells of theentorhinal cortex project to the dentate gyrus, and they terminate inthe outer two-thirds of the molecular layer of the dentate gyrus. Theaxons from layer III cells project bilaterally to the cornu ammonisareas CA1 and CA3 of the hippocampus, terminating in the stratumlacunose molecular layer.

In an illustrative embodiment, the administration is accomplished byinfusion of the LSD enzyme into one or both of the lateral ventricles ofa subject or patient. By infusing into the lateral ventricles, theenzyme is delivered to the site in the brain in which the greatestamount of CSF is produced. The enzyme may also be infused into more thanone ventricle of the brain. Treatment may consist of a single infusionper target site, or may be repeated. Multiple infusion/injection sitescan be used. For example, the ventricles into which the enzyme isadministered may include the lateral ventricles and the fourthventricle. In some embodiments, in addition to the first administrationsite, a composition containing the LSD enzyme is administered to anothersite which can be contralateral or ipsilateral to the firstadministration site. Injections/infusions can be single or multiple,unilateral or bilateral.

To deliver the solution or other composition containing the enzymespecifically to a particular region of the central nervous system, suchas to a particular ventricle, e.g., to the lateral ventricles or to thefourth ventricle of the brain, it may be administered by stereotaxicmicroinjection. For example, on the day of surgery, patients will havethe stereotaxic frame base fixed in place (screwed into the skull). Thebrain with stereotaxic frame base (MRI-compatible with fiduciarymarkings) will be imaged using high resolution MRI. The MRI images willthen be transferred to a computer that runs stereotaxic software. Aseries of coronal, sagittal and axial images will be used to determinethe target site of vector injection, and trajectory. The softwaredirectly translates the trajectory into 3-dimensional coordinatesappropriate for the stereotaxic frame. Burr holes are drilled above theentry site and the stereotaxic apparatus localized with the needleimplanted at the given depth. The enzyme solution in a pharmaceuticallyacceptable carrier will then be injected. Additional routes ofadministration may be used, e.g., superficial cortical application underdirect visualization, or other non-stereotaxic application.

One way for delivering a slow infusion is to use a pump. Such pumps arecommercially available, for example, from Alzet (Cupertino, Calif.) orMedtronic (Minneapolis, Minn.). The pump may be implantable. Anotherconvenient way to administer the enzymes is to use a cannula or acatheter. The cannula or catheter may be used for multipleadministrations separated in time. Cannulae and catheters can beimplanted stereotaxically. It is contemplated that multipleadministrations will be used to treat the typical patient with alysosomal storage disease. Catheters and pumps can be used separately orin combination.

The lysosomal storage diseases (LSD) includes over forty geneticdisorders, many of which involve genetic defects in various lysosomalhydrolases. Representative lysosomal storage diseases and the associateddefective enzymes are listed in Table 1.

Gaucher disease results as a consequence of an inherited deficiency ofthe lysosomal hydrolase glucocerebrosidase (GC), leading to theaccumulation of its substrate, glucosylceramide (GL-1), in the lysosomesof histiocytes. The progressive accumulation of GL-1 in tissuemacrophages (Gaucher cells) occurs in various tissues. The extent of theaccumulation is dependent in part on the genotype. Clinically, threedifferent Gaucher phenotypes are recognized, the non-neuropathic type 1,which is the most common with onset ranging from early childhood toadulthood, and the neuropathic types 2 and 3, presenting in infancy andearly childhood, respectively. Any of these phenotypes may be treated inaccordance with the present invention. The primary clinicalmanifestations common to all forms of Gaucher disease arehepatosplenomegaly, cytopenia, pathological bone fractures and,occasionally, pulmonary failure. A detailed discussion of Gaucherdisease may be found in the Online Metabolic & Molecular Bases ofInherited Diseases, Part 16, Chapter 146 and 146.1 (2007). In patientswith type 2 and type 3 Gaucher disease in whom there is significantcentral nervous system involvement, intraventricular delivery of thedefective LSD enzyme leads to improved metabolic status of the brain andpotentially the affected visceral (non-CNS) organs. Intraventriculardelivery of the defective LSD enzyme in subjects with Gaucher type 1disease leads to improved metabolic status of affected visceral(non-CNS) organs. There are animal models of Gaucher disease, which havederived from mouse models created by targeted disruption of thecorresponding mouse gene. For example, a Gaucher mouse model harboringthe D409V mutation in the mouse GC locus exists (Xu, Y-H et al. (2003).Am. J. Pathol. 163:2093-2101). The heterozygous mouse, gbaD409V/null,exhibits □5% of normal GC activity in visceral tissues and developslipid-engorged macrophages (Gaucher cells) in the liver, spleen, lungand bone marrow by 4 months of age. This model is a suitable system inwhich to evaluate the benefits of and to determine the conditions forthe intraventricular delivery of the defective LSD enzyme in subjectswith Gaucher disease.

Niemann-Pick disease (NPD) is a lysosomal storage disease and is aninherited neurometabolic disorder characterized by a genetic deficiencyin acid sphingomyelinase (ASM; sphingomyelin cholinephosphohydrolase, EC3.1.3.12). The lack of functional ASM protein results in theaccumulation of sphingomyelin substrate within the lysosomes of neuronsand glia throughout the brain. This leads to the formation of largenumbers of distended lysosomes in the perikaryon, which are a hallmarkfeature and the primary cellular phenotype of type A NPD. The presenceof distended lysosomes correlates with the loss of normal cellularfunction and a progressive neurodegenerative course that leads to deathof the affected individual in early childhood (The Metabolic andMolecular Bases of Inherited Diseases, eds. Scriver et al., McGraw-Hill,New York, 2001, pp. 3589-3610). Secondary cellular phenotypes (e.g.,additional metabolic abnormalities) are also associated with thisdisease, notably the high level accumulation of cholesterol in thelysosomal compartment. Sphingomyelin has strong affinity forcholesterol, which results in the sequestering of large amounts ofcholesterol in the lysosomes of ASMKO mice and human patients (Leventhalet al. (2001) J. Biol. Chem., 276:44976-44983; Slotte (1997) Subcell.Biochem., 28:277-293; and Viana et al. (1990) J. Med. Genet.,27:499-504.) A detailed discussion of NPD disease may be found in theOnline Metabolic & Molecular Bases of Inherited Diseases, Part 16,Chapter 144 (2007). There are animal models of NPD. For example, ASMKOmice are an accepted model of types A and B Niemann-Pick disease(Horinouchi et al. (1995) Nat. Genetics, 10:288-293; Jin et al. (2002)J. Clin. Invest., 109:1183-1191; and Otterbach (1995) Cell,81:1053-1061). Intraventricular delivery of the defective LSD enzymeleads to improved metabolic status of the brain and the affectedvisceral (non-CNS) organs.

Mucopolysaccharidoses (MPS) are a group of lysosomal storage disorderscaused by deficiencies of enzymes catalyzing the degradation ofglycosaminoglycans (mucopolysaccharides). There are 11 known enzymedeficiencies that give rise to 7 distinct MPS, including MPS I (Hurler,Scheie, and Hurler-Scheie Syndromes) and MPS II (Hunter Syndrome). AnyMPS can be treated in accordance with the present invention. A detaileddiscussion of MPS may be found in the Online Metabolic & Molecular Basesof Inherited Diseases, Part 16, Chapter 136 (2007). There are numerousanimal models of MPS, which have derived from naturally occurringmutations in dogs, cats, rats, mice, and goats, as well as mouse modelscreated by targeted disruption of the corresponding mouse gene. Thebiochemical and metabolic features of these animal models are generallyquite similar to those found in humans; however, the clinicalpresentations may be milder. For example, accepted models for MPS Iinclude a murine model [Clark, L A et al., Hum. Mol. Genet. (1997),6:503] and a canine model [Menon, K P et al., Genomics (1992), 14:763.For example, accepted models for MPS II include a mouse model [Muenzer,J. et al., (2002), Acta Paediatr. Suppl.; 91(439):98-9]. In the MPS thathave central nervous system involvement, such as is found in patientswith MPS I and MPS II, intraventricular delivery of the defective LSDenzyme leads to improved metabolic status of the brain and potentiallythe affected visceral (non-CNS) organs.

Pompe disease, or glycogen storage disease type II (GSDII), also termedacid maltase deficiency (AMD) is an inherited disorder of glycogenmetabolism resulting from defects in activity of the lysosomal hydrolaseacid alpha-glucosidase in all tissues of affected individuals. Theenzyme deficiency results in intralysosomal accumulation of glycogen ofnormal structure in numerous tissues. The accumulation is most marked incardiac and skeletal muscle and in hepatic tissues of infants with thegeneralized disorder. In late-onset GSDII, intralysosomal accumulationof glycogen is virtually limited to skeletal muscle and is of lessermagnitude. Electromyographic abnormalities suggestive of the diagnosisinclude pseudomyotonic discharges and irritability, but in juvenile- andadult-onset patients, the abnormalities can vary in different muscles.CAT scans can reveal the site(s) of affected muscles. Most patients haveelevated blood plasma levels of creatine kinase (CK) and elevations inhepatic enzymes, particularly in adult-onset patients, can be found.There are several naturally occurring animal models of the infantile-and late-onset disease. There is a knockout mouse model [Bijvoet A G etal., Hum. Mol. Genet. (1998); 7:53-62.]. Ameliorative effects of enzymetherapy have been described in knockout mice [Raben, N et al., Mol.Genet. Metab. (2003); 80:159-69] and in a quail model. Intraventriculardelivery of the defective LSD enzyme leads to improved metabolic statusof the brain and potentially the affected visceral (non-CNS) organs.

The neuronal ceroid lipofuscinoses (NCL) are a group ofneurodegenerative disorders distinguished from other neurodegenerativediseases by the accumulation of autofluorescent material (“agingpigment”) in the brain and other tissues. The major clinical featuresinclude seizures, psychomotor deterioration, blindness, and prematuredeath. Distinct subgroups of NCL have been recognized that differ in theage of onset of symptoms and the appearance of the storage material byelectron microscopy. Three major groups-infantile (INCL), classical lateinfantile (LINCL), and juvenile (JNCL, also referred to as Battendisease)—are caused by autosomal recessive mutations in the CLN1, CLN2,and CLN3 genes, respectively. The protein products of the CLN1(palmitoyl-protein thioesterase) and CLN2 (tripeptidyl peptidase orpepinase) genes are soluble lysosomal enzymes, whereas the CLN3 protein(battenin) is a lysosomal membrane protein, as is (tentatively) the CLN5protein. The identification of mutations in genes encoding lysosomalproteins in several forms of NCL has led to the recognition of thelipofuscinoses as true lysosomal storage disorders. Any subgroup of NCLcan be treated in accordance with the present invention. A detaileddiscussion of NCL disease may be found in the Online Metabolic &Molecular Bases of Inherited Diseases, Part 16, Chapter 154 (2007).Naturally occurring NCL disorders have been described in the sheep, dog,and mouse models have been derived by targeted disruption of acorresponding mouse gene [see e.g., Katz, M L et al., J. Neurosci. Res.(1999); 57:551-6; Cho, S K et al., Glycobiology (2005); 15:637-48.]Intraventricular delivery of the defective LSD enzyme leads to improvedmetabolic status of the brain and possibly the affected visceral(non-CNS) organs.

A detailed discussion of additional lysosomal storage disordersdisclosed in Table 1, in which intraventricular delivery of thedefective LSD enzyme in the disease, may be found in the OnlineMetabolic & Molecular Bases of Inherited Diseases, Part 16 (2007).

The above disclosure generally describes the present invention. Allreferences disclosed herein are expressly incorporated by reference. Amore complete understanding can be obtained by reference to thefollowing specific examples which are provided herein for purposes ofillustration only, and are not intended to limit the scope of theinvention.

Example 1

Animal Model

ASMKO mice are an accepted model of types A and B Niemann-Pick disease(Horinouchi et al. (1995) Nat. Genetics, 10:288-293; Jin et al. (2002)J. Clin. Invest., 109:1183-1191; and Otterbach (1995) Cell,81:1053-1061). Niemann-Pick disease (NPD) is classified as a lysosomalstorage disease and is an inherited neurometabolic disordercharacterized by a genetic deficiency in acid sphingomyelinase (ASM;sphingomyelin cholinephosphohydrolase, EC 3.1.3.12). The lack offunctional ASM protein results in the accumulation of sphingomyelinsubstrate within the lysosomes of neurons and glia throughout the brain.This leads to the formation of large numbers of distended lysosomes inthe perikaryon, which are a hallmark feature and the primary cellularphenotype of type A NPD. The presence of distended lysosomes correlateswith the loss of normal cellular function and a progressiveneurodegenerative course that leads to death of the affected individualin early childhood (The Metabolic and Molecular Bases of InheritedDiseases, eds. Scriver et al., McGraw-Hill, New York, 2001, pp.3589-3610). Secondary cellular phenotypes (e.g., additional metabolicabnormalities) are also associated with this disease, notably the highlevel accumulation of cholesterol in the lysosomal compartment.Sphingomyelin has strong affinity for cholesterol, which results in thesequestering of large amounts of cholesterol in the lysosomes of ASMKOmice and human patients (Leventhal et al. (2001) J. Biol. Chem.,276:44976-44983; Slotte (1997) Subcell. Biochem., 28:277-293; and Vianaet al. (1990) J. Med. Genet., 27:499-504.)

Example 2

“Continuous Intraventricular Infusion of rhASM in the ASMKO Mouse”

Goal: To determine what effect intraventricular infusion of recombinanthuman ASM (rhASM) has on storage pathology (i.e., sphingomyelin andcholesterol storage) in the ASMKO mouse brain

Methods: ASMKO mice were stereotaxically implanted with an indwellingguide cannula between 12 and 13 weeks of age. At 14 weeks of age micewere infused with 0.250 mg of hASM (n=5) over a 24 h period (˜0.01 mg/h)for four straight days (1 mg total was administered) using an infusionprobe (fits inside the guide cannula) which is connected to a pump.Lyophilized hASM was dissolved in artificial cerebral spinal fluid(aCSF) prior to infusion. Mice were sacrificed 3 days post infusion. Atsacrifice mice were overdosed with euthasol (>150 mg/kg) and thenperfused with PBS or 4% parformaldehyde. Brain, liver, lung and spleenwere removed and analyzed for sphingomyelin (SPM) levels. Brain tissuewas divided into 5 sections before SPM analysis (S1=front of brain,S5=back of the brain; see FIG. 1)

TABLE 2 Group Treatment n ASMKO .250 mg/24 h (1 mg total) 5 ASMKO None 4WT None 4

Results summary: Intraventricular infusion of hASM at 0.250 mg/24 h for4 continuous days (1 mg total) resulted in hASM staining and filipin(i.e., cholesterol storage) clearance throughout the ASMKO brain.Biochemical analysis showed that intraventricular infusion of hASM alsoled to a global reduction in SPM levels throughout the brain. SPM levelswere reduced to that of wild type (WT) levels. A significant reductionin SPM was also observed in the liver and spleen (a downward trend wasseen in the lung).

Example 3

“Intraventricular Delivery of hASM in ASMKO Mice II”

Goal: to determine lowest efficacious dose over a 6 h infusion period.

Methods: ASMKO mice were stereotaxically implanted with an indwellingguide cannula between 12 and 13 weeks of age. At 14 weeks of age micewere infused over a 6 hour period at one of the following doses of hASM:10 mg/kg (0.250 mg; n=12), 3 mg/kg (0.075 mg; n=7), 1 mg/kg (0.025 mg;n=7), 0.3 mg/kg (0.0075 mg; n=7), or aCSF (artificial cerebral spinalfluid; n=7). Two mice from each dose level were perfused with 4%parformaldehyde immediately following the 6 h infusion to assess enzymedistribution in the brain (blood was also collected from these todetermine serum hASM levels). The remaining mice from each group weresacrificed 1 week post infusion. Brain, liver, and lung tissue fromthese mice was analyzed for SPM levels as in study 05-0208.

TABLE 3 Group Treatment n ASMKO  0.250 mg (10 mg/kg) 12 ASMKO  0.075 mg(3 mg/kg) 7 ASMKO  0.025 mg (1 mg/kg) 7 ASMKO 0.0075 mg (.3 mg/kg) 7ASMKO aCSF 7 WT None 7

Results summary: Intraventricular hASM over a 6 h period led to asignificant reduction in SPM levels throughout the brain regardless ofdose. Brain SPM levels in mice treated with doses >0.025 mg were reducedto WT levels. Visceral organ SPM levels were also significantly reduced(but not to WT levels) in a dose dependent manner. In support of thisfinding hASM protein was also detected in the serum of ASMKO miceinfused with hASM protein. Histological analysis showed that hASMprotein was widely distributed throughout the brain (from S1 to S5)after intraventricular administration of hASM.

Example 4

“Intraventricular Infusion of rhASM in ASMKO Mice III”

Goal: To determine (1) the time it takes for SPM to reaccumulate withinthe brain (and spinal cord) after a 6 h infusion of hASM (dose=0.025mg); (2) if there are sex differences in response to intraventricularhASM administration (pervious experiments demonstrate that there are sexdifferences in substrate accumulation in the liver, whether or not thisoccurs in the brain is unknown).

Methods: ASMKO mice were stereotaxically implanted with an indwellingguide cannula between 12 and 13 weeks of age. At 14 weeks of age micewere infused over a 6 h period with 0.025 mg of hASM. Afterintraventricular delivery of hASM mice were sacrificed either at 1 weekpost infusion (n=7 males, 7 females), or at 2 weeks post infusion (n=7males, 7 females) or at 3 weeks post infusion (n=7 males, 7 females). Atsacrifice the brain, spinal cord, liver and lung were removed for SPManalysis.

TABLE 4 Group Treatment N Sacrifice male ASMKO .025 mg 7 1 week postinfusion Female ASMKO .025 mg 7 1 week post infusion male ASMKO .025 mg7 2 weeks post infusion Female ASMKO .025 mg 7 2 weeks post infusionmale ASMKO .025 mg 7 3 weeks post infusion Female ASMKO .025 mg 7 3weeks post infusion male ASMKO aCSF 7 1 week post infusion Female ASMKOaCSF 7 1 week post infusion male WT None 7 1 week post infusion FemaleWT None 7 1 week post infusion

Tissue samples are prepared for SPM analysis.

Example 5

“Effect of Intraventricular Infusion of rhASM on Cognitive Function inASMKO Mice”

Goal: to determine if intraventricular infusion of rhASM alleviatesdiseased induced cognitive deficits in ASMKO mice

Methods: ASMKO mice are stereotaxically implanted with an indwellingguide cannula between 9 and 10 weeks of age. At 13 weeks of age mice areinfused over a 6 h period with 0.025 mg of hASM. At 14 and 16 weeks ofage mice undergo cognitive testing using the Barnes maze.

Example 6

“hASM Protein Distribution within the ASMKO CNS after IntraventricularInfusion”

Goal: to determine the distribution of hASM protein (as function oftime) within the brain and spinal cord of ASMKO mice afterintraventricular infusion

Methods: ASMKO mice are stereotaxically implanted with an indwellingguide cannula between 12 and 13 weeks of age. At 14 weeks of age miceare infused over a 6 h period with 0.025 mg of hASM. Following infusionprocedure mice are either sacrificed immediately or 1 week or 2 weeks or3 weeks later.

TABLE 5 Indicates the infusion times that may be used with a particularenzyme for the treatment of the disease in which it is deficient asindicated in Table 1. INFUSION TIME more more more more more 1-5 5-1010-30 30-60 1-4 than than than than than Enzyme for Infusion BOLUS minmin min min hrs 4 hrs 5 hrs 6 hrs 7 hrs 8 hrsAspartylglucosaminidase•alpha•- X X X X X X X X X X X Galactosidase APalmitoyl Protein Thioesterase X X X X X X X X X X X TripeptidylPeptidase X X X X X X X X X X X Lysosomal Transmembrane Protein X X X XX X X X X X X Cysteine transporter X X X X X X X X X X X Acid ceramidaseX X X X X X X X X X X Acid•alpha•-L-fucosidase X X X X X X X X X X XProtective protein/cathepsin A X X X X X X X X X X XAcid•beta•-glucosidase, or X X X X X X X X X X XAcid•beta•-galactosidase Iduronate-2-sulfatase X X X X X X X X X X Xalpha•L-Iduronidase X X X X X X X X X X X Galactocerebrosidase X X X X XX X X X X X Acid•alpha•-mannosidase X X X X X X X X X X XAcid•beta•-mannosidase X X X X X X X X X X X Arylsulfatase B X X X X X XX X X X X Arylsulfatase A X X X X X X X X X X XN-Acetylgalactosamine-6-sulfate X X X X X X X X X X XAcid•beta•-galactosidase X X X X X X X X X X X N-Acetylglucosamine-1- XX X X X X X X X X X Acid sphingomyelinase X X X X X X X X X X X

REFERENCES

The disclosure of each reference cited is expressly incorporated herein.

-   1) Belichenko P V, Dickson P I, Passage M, Jungles S, Mobley W C,    Kakkis E D. Penetration, diffusion, and uptake of recombinant human    alpha-1-iduronidase after intraventricular injection into the rat    brain. Mol Genet Metab. 2005; 86(1-2):141-9.-   2) Kakkis E, McEntee M, Vogler C, Le S, Levy B, Belichenko P, Mobley    W, Dickson P, Hanson S, Passage M. Intrathecal enzyme replacement    therapy reduces lysosomal storage in the brain and meninges of the    canine model of MPS I. Mol Genet Metab. 2004; 83(1-2):163-74.-   3) Bembi B, Ciana G, Zanatta M, et al. Cerebrospinal-fluid infusion    of alglucerase in the treatment for acute neuronopathic Gaucher's    disease. Pediatr Res 1995; 38:A425.-   4) Lonser R R, Walbridge S, Murray G J, Aizenberg M R, Vortmeyer A    O, Aerts J M, Brady R O, Oldfield E H. Convection perfusion of    glucocerebrosidase for neuronopathic Gaucher's disease. Ann Neurol.    2005 April; 57(4):542-8.

1-41. (canceled)
 42. A kit for treating a patient with a lysosomalstorage disease which is caused by an enzyme deficiency, comprising: theenzyme; and a catheter, a pump, or a catheter and a pump for delivery ofsaid acid sphingomyelinase to the patient's brain ventricles, whereinthe kit is for administering a plurality of infusions of the enzyme tothe patient via intraventricular delivery to the brain in an amountsufficient to reduce said enzyme's substrate levels throughout saidbrain; wherein each infusion is performed at a rate such that theadministration of a single dose consumes more than two hours, andwherein the administration is such that at least 10% of the substratelevels throughout said brain of the patient are reduced.
 43. The kit ofclaim 42, wherein the lysosomal storage disease is Niemann-Pick A andthe enzyme is acid sphingomyelinase.
 44. The kit of claim 43, whereinthe acid sphingomyelinase is a human acid sphingomyelinase.
 45. The kitof claim 43, wherein the acid sphingomyelinase comprises an amino acidsequence with at least 95% amino acid sequence identity to an acidsphingomyelinase as shown in SEQ ID NO:1.
 46. The kit of claim 43,wherein the acid sphingomyelinase comprises an amino acid sequence asshown in SEQ ID NO:
 1. 47. The kit of claim 42, wherein: (a) thelysosomal storage disease is Mucopolysaccharidosis I syndrome and theenzyme is alpha-L-iduronidase, (b) the lysosomal storage disease isMucopolysaccharidosis II syndrome and the enzyme isiduronate-2-sulfatase, (c) the lysosomal storage disease is Gaucherdisease and the enzyme is glucocerebrosidase, (d) the lysosomal storagedisease is Pompe disease and the enzyme is alpha-glucosidase, (e) thelysosomal storage disease is classic late infantile Batten disease(CLN2) and the enzyme is tripeptidyl peptidase, or (f) wherein thelysosomal storage disease is Niemann-Pick B and the enzyme is acidsphingomyelinase.
 48. The kit of claim 42, wherein the rate is such thatthe administration of a single dose consumes more than four hours. 49.The kit of claim 42, wherein the rate is such that the administration ofa single dose consumes more than five hours.
 50. The kit of claim 42,wherein the rate is such that the administration of a single doseconsumes more than six hours.
 51. The kit of claim 42, wherein the rateis such that the administration of a single dose consumes more thanseven hours.
 52. The kit of claim 42, wherein the rate is such that theadministration of a single dose consumes more than eight hours.
 53. Thekit of claim 42, wherein the plurality of infusions is administered tomore than one ventricle of the brain.
 54. The kit of claim 53, whereinthe more than one ventricle of the brain includes the lateral ventriclesand the fourth ventricle.
 55. A kit for treating a patient with alysosomal storage disease which is caused by an enzyme deficiency,comprising: the enzyme; and a catheter, a pump, or a catheter and a pumpfor delivery of said acid sphingomyelinase to the patient's brainventricles, wherein the kit is for administering a single dose of theenzyme to the patient via intraventricular delivery to the brain in anamount sufficient to reduce said enzyme's substrate levels throughoutsaid brain; wherein the enzyme is administered at a rate such that theadministration of the single dose consumes more than two hours, andwherein the administration is such that at least 10% of the substratelevels throughout said brain of the patient are reduced.
 56. The kit ofclaim 55, wherein the lysosomal storage disease is Niemann-Pick A andthe enzyme is acid sphingomyelinase.
 57. The kit of claim 56, whereinthe acid sphingomyelinase is a human acid sphingomyelinase.
 58. The kitof claim 56, wherein the acid sphingomyelinase comprises an amino acidsequence with at least 95% amino acid sequence identity to an acidsphingomyelinase as shown in SEQ ID NO:1.
 59. The kit of claim 56,wherein the acid sphingomyelinase comprises an amino acid sequence asshown in SEQ ID NO:
 1. 60. The kit of claim 55, wherein: (a) thelysosomal storage disease is Mucopolysaccharidosis I syndrome and theenzyme is alpha-L-iduronidase, (b) the lysosomal storage disease isMucopolysaccharidosis II syndrome and the enzyme isiduronate-2-sulfatase, (c) the lysosomal storage disease is Gaucherdisease and the enzyme is glucocerebrosidase, (d) the lysosomal storagedisease is Pompe disease and the enzyme is alpha-glucosidase, (e) thelysosomal storage disease is classic late infantile Batten disease(CLN2) and the enzyme is tripeptidyl peptidase, or (f) wherein thelysosomal storage disease is Niemann-Pick B and the enzyme is acidsphingomyelinase.
 61. The kit of claim 55, wherein the enzyme isadministered at a rate such that the administration of the single doseconsumes more than four hours.
 62. The kit of claim 55, wherein theenzyme is administered at a rate such that the administration of thesingle dose consumes more than five hours.
 63. The kit of claim 55,wherein the enzyme is administered at a rate such that theadministration of the single dose consumes more than six hours.
 64. Thekit of claim 55, wherein the enzyme is administered at a rate such thatthe administration of the single dose consumes more than seven hours.65. The kit of claim 55, wherein the enzyme is administered at a ratesuch that the administration of the single dose consumes more than eighthours.